Citrullination in proteins of the polycomb repressor complex 1

Area of Science:

• Molecular Biology
• Epigenetics
• Enzymology

Background:

• Polycomb repressive complexes regulate gene expression via chromatin modification.
• Ring finger protein 1 (RING1) and RYBP are key components of these complexes.
• Peptidyl arginine iminohydrolases (PADIs) catalyze arginine to citrulline modification.

Purpose of the Study:

• To investigate the in vitro and in silico binding of PADI4 to specific peptides from RING1B and RYBP.
• To determine the effect of citrullination on the binding affinity between PADI4 and these peptides.

Main Methods:

• Far-UV circular dichroism (CD) and Nuclear Magnetic Resonance (NMR) spectroscopy to assess peptide structure.
• Fluorescence, biolayer interferometry (BLI), and isothermal titration calorimetry (ITC) to measure binding affinities.
• Molecular docking simulations to analyze peptide-enzyme interactions at the active site.

Main Results:

• Isolated peptides from RING1B and RYBP, in both wild-type and citrullinated forms, were monomeric and disordered.
• Peptide binding affinities for PADI4 were in the low micromolar range (Kd ~10-20 μM).
• Citrullination of peptides resulted in diminished binding affinity for PADI4.

Conclusions:

• The study demonstrates that isolated peptides of RING1B and RYBP can bind to PADI4.
• PADI4 targets the active site of the enzyme, interacting with catalytic residues.
• Arginine citrullination by PADI4 reduces the binding affinity of these peptides.